We propose to develop unique but related animal models of three regulatory genes in the interferon-inducible, doublestranded RNA- activated protein kinase (PKR) pathway. The models will be transgenic (Tg) mouse lines overexpressing PKR, as well as P58IPK (a cellular inhibitor of PKR), and the PKR phosphorylation substrate eIF-2alpha (we will use a dominant negative S51A mutant incapable of being phosphorylated by eIF-2alpha kinases). In vitro expression studies of PKR have established roles for the enzyme in the antiviral effects of interferons, the responses of uninfected cells to physiologic stresses, and in cell growth regulation. The possibility that PKR may function as a tumor suppressor and inducer of apoptosis suggests that this interferon-regulated protein kinase may be of central importance to the control of cell proliferation and transformation. The investigation of PKR in a complex biological system has not yet been fully explored. Tg mice will be generated by an embryonic stem (ES) cell culture method designed to decrease the number of mice currently needed when standard techniques are used. We will establish phenotypic characteristics to develop specific transgenic models for studies in a wide variety of scientific areas. The PKR overexpression Tg model is designed to increase phosphorylation of eIF-2alpha thus inhibiting initiation of protein translation and conserving vital cellular resources. This model would be applicable for studies in viral resistance (hepatitis, influenza, HIV), aging, tumor suppression by apoptosis, and NF-kB mediated cytokine patterns and signal transduction. The eIF-2alphaS51A overexpression Tg model is designed to decrease eIF-2alpha phosphorylation by eIF-2alpha kinases, thus allowing an increase in protein translation. This model would be applicable for studies in viral susceptibility (hepatitis and HIV), cellular proliferation, and aging. The p58IPK overexpression Tg model is designed to increase the inhibition of PKR activity, thus mimicking some of the features of the eIF-2alpha mice but affecting other substrates, such as NF-kB, in addition to eIF-2alpha. This model would be applicable for studies in viral resistance, stress response and heat shock protein pathways, and aging. These models have the potential for becoming well-used tools by the research community for investigating PKR regulatory pathway mechanisms, and will contribute to the knowledge of the biology of PKR systems across a wide and diverse array of disciplines.